A bioaffinity-based method for the determination of adrenaline and dopamine involving immobilized renatured DNA incorporated into the amperometric biosensor was developed. With the help of spectrophometry and voltammetry data, it was found that the composition of complexes formed with catecholamines and their stability constants were changing with concentration from [КАL 2 ] to [КА 2 L], where КА – catecholamine, L – base pare, and lg β eff = 10.1–13.5. The high affinity of adrenaline and dopamine to the immobilized renatured DNA molecule was revealed, thereby making it possible to run effective concentration of KA from the analyzed solutions of low concentration including multicomponent fluids. The minimum reporting level was found to be 5.5·10 –9 (adrenaline) and 2.0·10 –8 (dopamin) mol/L, respectively. The proposed method is characterized by high sensitivity, reproducibility (relative accuracy does not exceed 5%), and single determination time shorter than 30 min.

The area of existence of decavanadate ions, which lies within the OH-/V molar ratios of 0 to 0.5, was studied in the V2O5 - LiOH - H2O system. The increase in the OH-/V molar ratios from 0.5 to 1.0 leads to an increase in the ratio of metavanadate ions and then the ratios of pyro-and orthovanadate ions. The obtained lithium decavanadate solutions were successfully used for the synthesis of crystal nickel decavanadates Ni3V10O28 center dot 22H(2)O and a double nickel-cesium decavanadate Cs2Ni2V10O28 center dot 15H(2)O.

We carried out an investigation of seasonal and annual changes of the main water quality parameters (color and turbidity). Based on the revealed regularities, we established a correlation between those characteristics and the content of copper, zinc, iron and manganese ions in water and developed a costeffective and fast method for water ecological monitoring. The proposed method enables not only determination but also forecasting of the content of heavy metal ions in natural water.

Bioaffine methods were developed for determining indole-containing alkaloids ajmaline and vincristine. These methods are based on the proposed amperometric DNA-sensors and on immunoenzyme test-system with a spectrophotometric indication of the analytical signal. The complexing between ajmaline and immobilized native DNA (n-IDNA) allows effective preliminary concentration from test solutions on the biosensor. The time of analysis is 25-30 min, limit of detection (LOD) for ajmaline is 1.0 x 10(-10) M. The test-system utilized the immunological reaction of ajmaline with its antibodies and the enzyme marker, horseradish peroxidase, LOD is 1.5 x 10(-9) M. Anti-cancer vincristine interaction with immobilized renatured DNA (r-IDNA) was studied and determination was carried out using an amperometric DNA-sensor. Bioaffine membrane drug concentration and reactivation of the sensor was performed. The affinity binding constant K(bind) for vincristine-r-IDNA complex calculated by Scatchard's method was found to be high enough [(5.0 +/- 0.4) x 10(5) l/mol] confirming high specificity of the complexing with r-IDNA. The duration of the assay is 40 min. The developed method is characterized with an LOD of 1.1 x 10(-9) M, with the lack of the need for long sample processing. The pharmaceuticals were determined by those methods in model solutions of blood serum and in tablets and solutions for injections.

Bioaffine methods are developed for determining indole-containing alkaloid ajmaline, which has a cytostatic effect and is used as a cardiac drug. These methods are based on the proposed amperometric DNA-sensor and on immunoenzyme test system with the spectrophotometric indication of the analytical signal. The complex formation between ajmaline and immobilized native DNA allows ajmaline to be efficiently preconcentrated on the biosensor from test solutions. Optimum conditions for preconcentrating ajmaline and those for reactivating the biosensor for its repeated use are found. The time of analysis is 25–30 min, the determination limit for ajmaline is 3.0 × 10−10 M (RSD = 33%). In the test system, the immunological reaction of ajmaline with its antibodies and the enzyme marker, horseradish peroxidase, are used. The determination limit is 4.0 × 10−9 M (RSD = 33%). Ajmaline is determined by the two methods in model solutions of blood serum and in tablets and solutions for injections.

The interaction of alkaloids with DNA immobilized into amperometric biosensor has been studied taking ajmaline as an example. The alkaloid can be effectively concentrated from analytes due to its complex-forming with native immobilized DNA. The optimal conditions for concentrating and reactivation of the sensor have been found out. The method based on the developed DNA-sensor has been proposed for the determination of indole alkaloid ajmaline used as a cardiological medicine and having cytostatic action. The method is characterized by sensitivity, good reproducibility and speed.

The analytical capabilities of electrochemical biosensors based on nucleic acids are systematized. Immobilization methods that retain the biological activity of nucleic acids and provide an opportunity to use them as multipurpose analytical reagents are described. The use of the above sensors in bioaffinity assays for determining DNA and its effectors in biochemical analysis and environmental monitoring and for determining the nucleotide composition of DNA is demonstrated in many examples.

Amperometric biosensors were developed on the basis of stationary mercury-film and glassy-carbon electrodes and DNA or its fragments, oligodeoxynucleotides (ODNs), immobilized in a nitrocellulose matrix. Taking into account the high affinity of Cu(II) and Fe(III) ions to denatured DNA ((19.1 ± 0.1) ×105 and (1.4 ± 0.3) × 105 L/mol, respectively), a procedure was proposed for the voltammetric determination of these ions in natural materials and blood serum at a level of n × 10−11 M. This procedure involves analyte pre-concentration on a DNA-containing biosensor. An ODN-containing biosensor (DNA probe) was used in the study of DNA hybridization for the highly specific determination of its nucleotide sequence.

The DNA-modified membrane electrode was prepared by casting a mixture of nitrocellulose (NC) with target DNA (tDNA) in organic solvent on glassy carbon electrode (GCE). Unlabeled polymerase chain reaction (PCR)-amplified human genomic sequence (628 bp) or synthetic oligodeoxynucleotides (ODNs) were used as tDNAs, creating a recognition layer. Biotinylated ODNs were used as hybridization probes to recognize specific nucleotide sequences. The hybridization events were detected via an enzyme-linked electrochemical assay involving binding of streptavidin-coupled alkaline phosphatase (SALP) to the biotin labels of the probe bound to tDNA. After the probe hybridization and SALP binding, the electrode was immersed into an electroinactive enzyme substrate (1-naphthyl phosphate). The alkaline phosphatase converted the inactive substrate into electroactive 1-naphthol that penetrated through the NC membrane to the GCE surface and was subsequently detected using an anodic voltammetric signal. The optimized method offered a good discrimination between complementary and nonspecific DNAs and yielded well-defined responses for both single-copy and repetitive tDNA sequences. In contrast to previously published methods using electrodes with mechanically attached membranes, the previously mentioned electrode is easily amenable to parallel DNA analysis.

The immobilized single-stranded DNA (ssIDNA) has been found to be a very effective biospecific analytical reagent when used in a newly developed bioaffinity method of the determination of heavy metals based on the amperometric DNA-based biosensor. This has been concluded from the comparative study of the complexing of heavy metals with double-stranded DNA, single-stranded DNA, and ssIDNA, using Fe(III) and Cu(II) as a model (metal/nucleotide ratio and stability constants are maximum for ssIDNA), from the study of adsorption of Fe(III), Cu(II), Pb(II), and Cd(II) on nitrocellulose membranes, containing single-stranded DNA, and from the determination of their binding constants with ssIDNA. According to these data, the chosen heavy metals can be lined up in a series of binding strengths with ssIDNA: Cu(II) > Pb(II) > Fe(III) > Cd(II). The method of the determination of heavy metals is based on biospecific preconcentration of metal ions on the biosensor followed by the destruction of DNA-metal complexes with ethylenediaminetetraacetate and voltammogram recording has been proposed. The lower detection limits are 4.0 x 10(-11), 1.0 x 10(-10), 1.0 x 10(-9), and 5.0 x 10(-9) M for Cu(II), Pb(II), Cd(II), and Fe(III), respectively. The heavy metals have been assayed in multicomponent environmental and biological systems such as natural and drinking water, milk, and blood serum samples.

Amperometric biosensor (BS) has been elaborated based on the stationary mercury-film electrode (SMFE) with silver support and cellulose nitrate (CN) membrane containing immobilized single-stranded DNA (ssIDNA). The sorption isotherms and ssDNA–heavy metal binding constants have been obtained with the BS. According to these data, the chosen heavy metals form the following series of binding strength with ssIDNA: Pb(II)>Fe(III)>Cd(II). It has been found that upon the competitive adsorption, there exists practically simultaneous sorption of different ions at ssIDNA containing membrane. The method of the determination of heavy metals based on preconcentration of metal ions on the BS followed by the destruction of DNA–metal complexes with ethylenediamine tetraacetate (EDTA) and voltammogram recording has been proposed. The lower limits of detectable contents are 1.0×10−10, 1.0×10−9 and 1.0×10−7 mol l−1 for Pb(II), Cd(II) and Fe(III), respectively. Heavy metals have been assayed in natural and drinking water, milk and blood serum samples even under simultaneous presence with a selectivity factor of 1:10. The effect of matrix components has been estimated.

A method of denatured DNA immobilisation on cellulose nitrate film has been developed. A modified film of uniform and stable surface has been used as a bio-sensitive part of amperometric DNA biosensor based on the stationary mercury-film covered silver electrode. The biosensor has been used to devise a new variant of solid-phase immunoassay of auto-antibodies (Ab) in blood serum without separation of components. The content of auto-Ab was monitored by measuring the currents of catalytic hydrogen evolution (with potentials of −1.2 and −1.4 V) resulting from the complexing of Pt(II) with DNA or auto-Ab respectively. The determination has been performed within a wide concentration area of 5.0×10−10 to 7.0×10−8 M. The limit of detection is 3.0×10−10 M. The affinity constants for the immunoreaction of DNA–antibodies have been found to be 1.25×109 and 2.50×108 M−1, which confirms the specificity of the interaction. The protocol of the immunoassay has been proposed and the procedure of diagnosing Aleutian mink disease (AMD) has been described here.

The sorption of Pb(II) and Cd(II) ions on a nitrocellulose membrane modified with denaturated deoxyribonucleic acid (d-DNA) was studied. A method for constructing isotherms of adsorption of heavy metals on the surface of an amperometric biosensor incorporating a d-DNA-modified nitrocellulose membrane and a stationary mercury-film electrode was proposed. An analysis of isotherms of joint sorption of Pb(II) and Cd(II) from a mixture of these ions showed that Pb(II) is bound to d-DNA more strongly than Cd(II), in agreement with the theoretical assumption that there are additional modes for binding Pb(II) with nitrogen-containing heterocycles of the d-DNA molecule. The constants of binding of Pb(II) and Cd(II) on d-DNA were determined by the Scatchard method.

A highly sensitive amperometric biosensor for selective determination of platinum in biological materials, pharmaceuticals, and other media is described. The biosensor is designed on the basis of a denatured deoxyribonucleic acid (DNA) immobilized on a nitrocellulose membrane and allows Pt(IV) and Pt(II) to be analyzed in the concentration ranges of 1.0 x 10(-6) 0.5 x 10(-10) M and 1.0 x 10(-6)-0.5 x 10(-8) M, respectively. The method features good reproducibility and simplicity of sample preparation, conditions for reactivation of the biosensitive component of the biosensor for multiple reuse are specified.

A new method for the immobilization of DNA on a nitrocellulose membrane was proposed. The activity of immobilized biomolecules is retained their activity. The immobilized DNA was used as the biosensitive part of an amperometric sensor based on a stationary mercury-film electrode. To extend the analytical capabilities of the sensor, the peak current of catalytic hydrogen evolution at -1.4 V was examined (this peak current occurs in the complexation of platinum(II) with antibodies to DNA). The biosensor was used for determining antibodies to DNA in the concentration range 5.0 x 10(-10)-7.0 x 10(-8) M. An immunoassay procedure was proposed.

New fast way of DNA immobilization on cellulose nitrate membrane which retains maximum of DNA activity has been developed. The amperometric sensor based on the membrane obtained and stationary mercury-film covered silver electrode was used for the determination of the DNA specific antibodies in blood serum. The antibodies content was monitored by measuring the hydrogen catalytic liberation peak current at the potential of -1.2 V which was resulted from Pt(II) complexing with DNA. The scheme of this process was proposed. The peak current value at -1.2 V decreases with an increase of antibodies concentration in blood serum due to partial shielding of the DNA-Pt(II) catalytically active complexes through biospecific interaction of DNA with antibodies. The Ab can be determined in the concentration area of 1.2x10(-9) - 5.0x10(-10) M. The sensor can be used within a month without loss of activity.

A biochemical sensor based on a stationary mercury film electrode and a cellulose nitrate film containing either DNA molecules or molecules of antibodies to DNA was developed to determine the concentration of DNA (antibodies to DNA) and to diagnose autoimmune diseases. The peak currents of catalytic hydrogen liberation upon the formation of Pt(II)-DNA (antibodies to DNA) complexes were used as the analytical signal. The detection limits were 0.075 and 10 mu g/mL for antibodies and DNA, respectively.

A biochemical biosensor was developed on the basis of a steady-state Hg-film electrode and either DNA molecules or antibodies to DNA immobilized in a cellulose nitrate film. This biosensor is designed to measure the concentration of DNA (or antibodies to DNA), and it can be used in the diagnosis of autoimmune diseases. Electric current of catalytic H-2 evolution caused by complexing between DNA (antibodies to DNA) and Pt (II) was used as an analytical signal. The detection sensitivity threshold for DNA and antibodies to DNA was 10 and 0.075 mu g/ml, respectively.

A method of DNA immobilization on cellulose nitrate films has been developed. Modified films of uniform and stable surface have been used to devise two variants of solid-phase enzyme immunoassays of antibodies. The coimmobilization of enzyme label (cholinesterase) and the DNA molecules makes it possible to carry out the procedure of solid-phase enzyme immunoassay without any separation of components. Thus, it takes only 15 min to diagnose an autoimmune disease (Aleutian disease of minks) with the immunoenzyme amperometric sensor, with a lower detection limit for antibodies of 0.5 x 10(-10) M. For scaled diagnosing, solid-phase enzyme immunoassay on DNA-modified films with prior separation of components and spectrophotometric registration of peroxidase activity has been developed, The time for determination was 30 min, with a lower detection limit of 7.4 x 10(-12) M.